Cancer results from uncontrolled cellular growth, due to a breakdown of normal cellular responses and cell cycle pathways, characterized by the progressive accumulation of lesions in the tumor genome. The number, severity and types of these lesions determine the biological properties of a given tumor. Genomic rearrangements, often the result of a translocation, interstitial deletion, or chromosomal inversion, account for the onset, development and progression of many tumorigenic diseases and predispositions to such diseases. Oncogenic fusion genes may lead to a gene product with a new or different function from the two fusion partners or upregulation of the gene product. Most fusion genes are found from hematological cancers, sarcomas and prostate cancer (Mitelman F, et al., “The impact of translocations and gene fusions on cancer causation.” Nat Rev Cancer. 2007 April; 7(4):233-45. Epub 2007 Mar. 15; Teixeira M R, “Recurrent fusion oncogenes in carcinomas.” Crit Rev Oncog. 2006 December; 12(3-4):257-71) and lymphomas (Vega F, Medeiros L J, “Chromosomal translocations involved in non-Hodgkin lymphomas.” Arch Pathol Lab Med. 2003 September; 127(9): 1148-60).
Most if not all fusion genes have been discovered as a result of pursuing to the specific goal of isolating a fusion gene expected to be associated with a chromosomal translocation, with one recent exception representing a more general approach (Raphael B J, et al. “A sequence-based survey of the complex structural organization of tumor genomes.” Genome Biol. 2008; 9(3):R59. Epub 2008 Mar. 25). Karyotypic detection of translocations has been very useful for cancer researchers, especially in identification of chromosomal translocation. Clinically, the presence or absence of specific translocations has therapeutic and prognostic implications. More fundamentally, genes identified at the translocation breakpoints are strong candidates for involvement in malignant transformation (Sanchez-Garcia, I. Anna. Rev. Genet. 1997 40 31:429-453). These translocations serve as markers of the malignant state and can be either the cause or the consequence of the transformed state. For example, the Philadelphia chromosome is a specific t(9;22)(q34;q11) translocation that fuses the B-cell antigen receptor gene BCR and the 45 ABL oncogene (De Klein A, et al. “A cellular oncogene is translocated to the Philadelphia chromosome in chronic myelocytic leukaemia.” Nature. 1982 Dec. 23; 300(5894):765-7). This fusion is thought to represent the crucial event in the development of chronic granulocytic leukemia. However, this translocation can also appear later in the course of multiple forms of leukemia. Solid tumors as well may have characteristic translocations, suggesting that the development of an unstable chromosomal state increases the likelihood of translocations which in turn 55 increase the likelihood of tumor progression (Rabbitts T H, “Chromosomal translocations in human cancer.” Nature 1994 Nov. 10; 372(6502):143-9; Sanchez-Garcia I, “Consequences of chromosomal abnormalities in tumor development.” Annu Rev Genet. 1997; 31:429-53).
Chromosomal rearrangements are involved in a multiplicity of cellular events, which includes oncogenesis. The rearrangements can be detected microscopically, but determining the segments of DNA actually involved in the rearrangement is a tedious process, especially within a large DNA region. There are as many as 50,000 of these rearrangements with almost all having essentially unknown fusion points, i.e. the precise DNA segments are still undetermined. These rearrangements are listed in what is called the Mitelman database at an NIH web site, which contains over 50,000 case reports all representing likely unique chromosomal rearrangements. In all cases they are associated with some human pathological condition, mostly cancer. In most cases, limited DNA segments defining the novel chromosome junctions are not known.
The study of cancer fusion genes, arising from chromosomal translocations during cancer cell development, has led to a much more sophisticated understanding of the basis of cancer and to designer drugs specifically targeted to certain cancers. For example, translocations that have fused the c-myc gene with the IgH gene have led to the understanding that part of the development of Burkitt's lymphoma is due to an abnormal and apparently perennial activation of the c-myc gene, which in turn stimulates cell proliferation (Taub, R., et al. “Translocation of the c-myc gene into the immunoglobulin heavy chain locus in human Burkitt lymphoma and murine plasmacytoma cells.” Proc. Nat'l Acad. Sci. USA. 1982 December; 79(24):7837-41; Dalla-Favera, R., et al. Human c-myc one gene is located on the region of chromosome 8 that is translocated in Burkitt lymphoma cells. Proc. Nat'l Acad. Sci. USA. 1982 December; 79(24):7824-7; Neel, B. G., et al. Two human c-onc genes are located on the long arm of chromosome 8. Proc. Nat'l Acad. Sci. USA. 1982 December; 79(24):7842-6) and the understanding of the structure of the bcr-abl protein, resulting from the fusion of the bcr and abl genes (Heisterkamp, N., et al. Structural organization of the bcr gene and its role in the Ph′ translocation. Nature. 1985 Jun. 27-Jul. 3; 315(6022):758-61), led to the discovery and use of Gleevec (Buchdunger, E., et al. Inhibition of the Abl protein-tyrosine kinase in vitro and in vivo by a 2-phenylaminopyrimidine derivative. Cancer Res. 1996 Jan. 1; 56(1):100-4; Druker, B. J., et al. Effects of a selective inhibitor of the Abl tyrosine kinase on the growth of Bcr-Abl positive cells. Nat. Med. 1996 May; 2(5):561-6), which efficiently retards the progress of chronic myelocytic leukaemia without the side of effects of less specific, anti-proliferative drugs. Moreover, it is expected that fusion proteins may be used to generate cancer specific immune responses (Chiarle, R., et al. The anaplastic lymphoma kinase is an effective oncoantigen for lymphoma vaccination. Nat. Med. 2008 June; 14(6):676-80. Epub 2008 May 11).
Currently, chromosomal fusion cancer testing requires laboratory personnel to identify the point of chromosomal fusion by karyotype analysis, followed by fluorescence in-situ hybridization (FISH) analysis of BAC clones to identify DNA region involved. Finally, the genes, or regions, in proximity to the BAC clone are analyzed for a fusion event using mRNA-based assays, such as reverse PCR. Alternatively, a BAC library is created from cells possessing a translocation. The end sequences of these clones are sequenced and analyzed by computer to identify any two “end sequences” that are not on the same chromosome
A precise diagnosis is the first requirement for rational therapy, since each individual patient, as well as each individual tumor, has certain unique genetic traits. These differences in patients and tumors with similar phenotypic characteristics may not have the same underlying genotypes, and therefore, may respond differently to the same treatment. The classical histopathological and clinical criteria used to assess the likelihood of response to the most commonly used modalities used to treat cancer and other diseases and disorders are inadequate predictors of treatment efficacy. A case-by-case approach to identifying fusion genes is inefficient, with over 50,000 reported disease-associated chromosomal rearrangements in the Mitelman database. Consequently, there is a significant and unmet need for accurate diagnostic methods that improve patient care and disease outcome.